The XVIII Biennial International Workshop on the Smuts and ...

The XVIII BiennialInternational Workshopon the Smuts and Bunts

February 3 – 5, 2014Tune Kursuscenter near Copenhagen, Denmark

Organized bySøren K. Rasmussen (University of Copenhagen),Philipp Steffan (University of Copenhagen) and

Anders Borgen (Agrologica)

Scientific Advisory BoardHermann Bürstmayr (BOKU Vienna, Austria)Bent J. Nielsen (Aarhus University, Denmark)

Gunter Backes (University of Kassel, Germany)Anders Borgen (Agrologica Consulting, Mariager, Denmark)Søren K. Rasmussen (University of Copenhagen, Denmark)

Veronika Dumalasová (Crop Research Institute, Prague, Czech Republic)Denis Gaudet (Agriculture and Agri-Food, Canada)

The XVIII Biennial International Workshop on

the Smuts and Bunts

February 3rd to 5th 2014 at the Tune Kursuscenter, Denmark.

http://plen.ku.dk/english/research/plant_soil/breeding/conference-smuts-and-bunts/

Final Programme

Monday – February 3, 2014

14.00 – 15.30 Registration, rooms, coffee/ tea and cake 15.30 – 15.35 Welcome by Søren K. Rasmussen Session 1 (Chair Søren K. Rasmussen) 15:35 – 16.15 Keynote talk 1:

What don't we (I) know about smuts and bunts, by Blair Goates 16:15 – 16:45 Experiences and problems with loose and covered smut in organic barley breeding, by

Karl Josef Müller 16:45 - 17:00 Break 17:00 - 17:10 Does Thule III have resistance gene Bt13? By Anders Borgen 17:10 - 17:20 Can saponins control bunts and smuts ? two pilot studies by Ole Søgaard Lund 17:20 - 17:50 Effect of essential oils on germination of Tilletia caries and T. controversa teliospores,

Pavel Rysanek 17:50 - 18:20 Genome-wide association study for common bunt resistance in wheat and Creation of

Common Bunt resistant Composite Cross Populations, by Philipp Steffan 18:30-19:30 Diner 20:00- Coffee, tea; fruits and snacks

Tuesday – February 4, 2014

07:45 – 09:00 Breakfast, buffet Session 2 (Chair Anders Borgen) 09:00 – 09:40 Keynote talk 2:

Pathogen evolution and loss of effective host resistance genes? Common bunt, a case study, by Denis A. Gaudet

09:40 – 10:10 International winter wheat improvement program: breeding modern germplasm, landraces and synthetics for disease resistance including common bunt, by Alexey Morgunov

10:10 – 10:40 Coffee, tea 10:40 – 11:10 Mapping of Common Bunt Resistance Gene bt9 in hexaploid wheat, by Philipp Steffan 11:10 – 11:40 Outcome of the European Tilletia Ringtest, by Fabio Mascher 12:00 – 13:30 Lunch buffet

Tuesday – February 4, 2014

Session 3 (Chair Gunter Backes) 13:30 – 14:10 Fifty years of spores: breeding for resistance to dwarf bunt of wheat (Tilletia

contraversa Kühn) in the Western US, by David Hole 14:10 – 14:40 Screening for resistance to common bunt (Tilletia tritici) in Danish spring and winter

varieties of wheat and triticale, by Bent Nielsen 14:40 – 15:10 The effect of spore origin when screening for resistance against common bunt, by

Anders Borgen 15:10 – 15:40 Coffee, tea and cake 15:40 – 16:10 Common bunt and dwarf bunt resistance in winter wheat cultivars, by Veronika

Dumalasova 16:10 – 16:50 Breeding for organic agriculture - Dwarf bunt resistance in winter wheat, by Almuth

Elise Müllner Poster presentation

Study on time duration of the infestation potential of common bunt (Tilletia caries) and dwarf bunt (T. controversa) spores of wheat in soil and farmyard manures taking into account different crop rotation systems in ecological farming, by Robert Bauer Phosphatases from Ustilago maydis and possible role during infection, by Søren K. Rasmussen

18:00 - 20:00 Workshop diner, 3 course 20:00 - Coffee, tea; fruits and snacks

Wednesday – February 5, 2014

07:45 – 09:00 Breakfast, buffet Session 4 (Chair Veronika Dumalasova) 09:00 – 09:30 Common bunt caused by Tilletia caries: Evaluation of viable spores, set up of a protocol

to access transmission to plantlets and damage threshold, by Geoffrey Orgeur 09:30 – 10:00 New sources of resistance among landraces in the USDA National Small Grains

Collection, by Blair Goates 10:00 – 10:30 Coffee, tea 10:30 – 11:00 Why Organic Agriculture doesn’t want to use fungicides, by Gunter Backes 11:00 – 11:45 Proceedings: A special issue in a dedicated journal based on presentations at the

workshop. Are you interested and do you have the time to prepare manuscript? Who will host the XIX Workshop? Closing the workshop. v. Søren K. Rasmussen

12:00 – 13:00 Lunch buffet 13:00 – Departure

Wednesday 13:30 – 15:30

Roskilde Cathedral: Guided tour in English 14:00-15:30

Please, sign up if you would like to join this guided tour in the Cathedral on your way back.

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The XVIII Biennial International Workshop on the Smuts and Bunts Copenhagen, Feb 3–5, 2014

Experiences and Problems with Loose and Covered Smut inOrganic Barley Breeding

Karl-JosefMuller

Cereal Breeding Research Darzau, Darzau Hof 1, 29490 Neu Darchau, [email protected]

For organic seed multiplication loose and covered smuts are a challenge as no organic seedtreatment is sufficient. Since more than ten years there is experience and more or less successwith using different sources of resistances in breeding at Cereal Breeding Research Darzau.The system of spreading loose smut in the trials will be presented and some phenomenons andopen questions on how to deal with diversity in both - smuts and sources of resistence - will beexplained.


Does Thule III Have Resistance Gene Bt13?

Anders Borgen

Agrologica, Houvej 55, 9550 Mariager, [email protected]

All over the world, Thule III is used as differential variety for the resistance gene Bt-13against common bunt. It can therefore be strange that I can question whether Thule III haveBt-13, so let me elaborate the question. I also use Thule III as a standard for Bt-13, and I gotthis variety from GRIN PI 181463. This variety does indeed have a major gene against commonbunt and since we all use it as a standard for Bt-13 this gene is per definition Bt-13. What stillpuzzles me is that this variety does not look like Thule III. It says in the GRIN database thatit is of Swedish origin with the pedigree Thule II/Sammet. PI 181463 is a short, lodging andweak variety with a very poor winter hardiness, and it does not at all look like a Swedish winterwheat. I therefore ordered Thule III (NGB6714) and Thule II and Sammet from NordGen, andthey are all very different from PI 181463. The varieties show different resistance pattern to thecommon bunt than PI-181463. The question is therefore not whether PI 181463 carry Bt-13, butwhether PI 181463 is really Thule III?


Effect of Treating Seeds with an Agave Extract in order to ControlCommon Bunt in Wheat and Loose Smut in Barley

Anders Borgen1, Marianne Andresen2, Ernest Rashid Mbega2, and Ole Søgaard Lund2,*

1Agrologica, Houvej 55, 9550 Mariager, Denmark2University of Copenhagen, Department of Plant and Environmental Sciences, Thorvaldsensvej 40, 1871 Frederiksberg,

Denmark*[email protected]

Aqueous extract from the sisal agave, A. sisalana, is known to contain antifungal activity. Intwo small field experiments we have tested the effect of seed treatment using A. sisalana extractin order to control common bunt, Tilletia caries, in winter wheat and loose smut, Ustilago nuda,in spring barley, respectively. Disease incidence of plants from treated seeds was compared toincidences in plants from non-treated seeds and seeds treated with pure water. In comparisonto controls, a reduction of disease incidence of T. caries by 75% was observed in winter wheatand a reduction of U. nuda incidence by 20% was observed in spring barley following seedtreatment with A. sisalana extract. The observed difference in efficiency towards these twopathogens is likely to reflect that spores of T. caries are located on the seed surface and thereby immediately exposed to this type of antifungal seed treatment whereas the seed-borne stageof U. nuda is located in or near the embryo of the seed, and thereby better protected againstantifungal molecules applied from the outside to the seed.


Effect of Essential Oils on Germination of Tilletia caries and T.controversa Teliospores

Pavel Rysanek

Plant Protection, Czech University of Life Sciences, Kamycka 129, 16521 Prague, Czech [email protected]

Bunts caused by Tilletia caries and T. controversa are important diseases of wheat. Causalfungi are seed- and/or soilborne. The protection fully depends on seed treatment and/or croprotation. Several fungicides may be used in conventional agriculture, but their use is excludedin ecological agriculture. In the past, some essential oils were proved efficient in the controlof some pests and pathogens. So, we also tested their effect on germination of T. caries and T.controversa teliospores in vitro. A range of essential oil concentrations from 0.05 µl to 0.8 µl/10ml of water agar was tested. Fungicides “Dividend” and “Celest” and also untreated agar wereused as controls. Teliospores were sown onto treated agar surface. Petri dishes were placedinto the thermostat at 16 °C and 5 °C + light for T. caries and T. controversa, respectively.Germinated spores were counted during 3rd, 5th, 7th and 10th day in the case of T. caries and35th, 42nd and 49th day in the case of T. controversa. Altogether 34 essential oils were tested.The highest concentration of all of them had effectiveness comparable to commercial fungicidesand some of them were still quite well effective even at the lowest concentration. The mosteffective oils were also tested for wheat seed treatment in pot experiments. The presence of T.caries in plants was checked by PCR two months after sowing. Some formulations completelyprevented plant infection. Thus, the use of certain essential oils is promising relatively safe wayof bunt control.


Genome Wide Association Study for Common Bunt Resistance inWheat and Creation of Common Bunt Resistant Composite Cross

Populations

Philipp Steffan1, Gunter Backes1,2, Søren K. Rasmussen1, and Anders Borgen3,*

1University of Copenhagen, Department of Plant and Environmental Sciences, Thorvaldsensvej 40, 1871 Frederiksberg,Denmark

2University of Kassel, Department of Organic Agricultural Sciences, Nordbahnhofstrasse 1a, 37213 Witzenhausen, Germany3Agrologica, Houvej 55, 9550 Mariager, Denmark

*[email protected]

Common bunt remains a major challenge to organic wheat production in Denmark. In orderto increase our understanding of the genetic components involved in common bunt resistancea genome wide association study involving 250 wheat lines was conducted. Lines were grownin two replications per year in Mariager, Denmark. Infection was recorded as the percentageinfected ears of all ears per line and replication.

Wheat lines were genotyped with DArT markers (Diversity Arrays Technology, Australia),yielding 1824 polymorphic marker. A compressed mixed linear model accounting for bothpopulation structure and familial relatedness was used to estimate marker effects on commonbunt resistance score. In total, 3 QTL for common bunt resistance could be identified, 1 QTLlocated on chromosome 2B and the others on chromosome 7A.

Wheat composite cross populations are created by the inter crossing of varieties and the sub-sequent bulking of the offspring, creating highly diverse populations with an increased resiliencetowards environmental impacts. Such a buffering is anticipated to be of great benefit in organicfarming systems with a reduced impact of agrochemicals.

A number of 22 winter wheat varieties with different degrees of common bunt resistances wasused in 33 crosses. The offspring was bulked in two different ways: ’Population 1’ was createdby bulking equal amounts of F2 seeds from each cross. In order to build the second population160 head rows, from seed of the F2 of the crosses, with less than 2% infection were selectedto form “Population shr” (selected hear rows). Both populations were grown both under heavycommon bunt inoculum pressure in order to select for resistance among its plants, and withoutcommon bunt disease pressure as a control. Disease incidence was recorded in both populationsin generations F4 and F5, and the populations grown without disease pressure were compared totheir parents in a two location yield trial.

Infection levels between “Population 1” and “Population shr” differed significantly with dis-ease incidences of 14% and 4% for “Population 1” and “Population shr”, respectively. In a twolocation yield trial the population yield did not differ significantly from the mean parental yield.We believe that it is possible to use such wheat composite cross populations as a germplasmsource for organic breeding, and it should also be possible to give improved populations directlyto farmers for commercial wheat production. The resistance development in the populations willbe continued for 3 more years.


Pathogen Evolution and Loss of Effective Host Resistance Genes:Common Bunt, a Case Study

D.A. Gaudet

AAFC Lethbridge Research Center, Box 3000 Lethbridge Alberta, Canada T1J [email protected]

Common bunt has historically been an important disease of wheat but current modern agri-cultural production systems that utilize both fungicides and genetic resistance for control has ledto the virtual disappearance of the pathogen in most western countries. In the Pacific Northwest(PNW) of the U.S.A., resistance (R) gene deployment beginning in the1930’s was ineffectivefor controlling bunt since newly deployed R-genes were soon defeated by virulent races. Anexcellent recounting of the history of the loss of resistance to common bunt in wheat varietieshas been detailed by Kendrick and Holton. This history clearly demonstrated that virulencechanges in common bunt races in the PNW of the USA occurred in advance of the release andwidespread deployment of individual R-genes. The situation whereby a large proportion of thewheat acreage in the PNW from 1900–1950s was seeded to bunt-susceptible cultivars with R-gene resistant cultivars being grown on limited acreages, permitted many cycles of mutationand recombination within the bunt population over the region that generated the necessary viru-lence. In some regions of the developing world where periodic outbreaks of bunt occur in wheat,races possessing high and complex virulence have been identified. In Canada, the deploymentof the ‘Hope’ gene for bunt resistance became widespread in the 1950s and remains effectivein many current varieties. Bt10 was introduced into breeding programs to counter high levelsof susceptibility which coincided with the introduction of the semi-dwarfing genes (Rht) genesin the 1980’s; Rht genes were associated with high susceptibility to many diseases includingcommon bunt. Resistance to common bunt remains mandatory in new wheat varieties registeredfor western Canada.

Organic wheat production practices that disallow fungicide use, is again leading to a reemer-gence of bunt in wheat and increased inoculum levels in the environment. In order to reducerate of evolution of common bunt, a strategy to stop inoculum production must be coordinatedand implemented. This must involve active R-gene deployment, pyramiding of two or moreeffective genes coupled with judicious fungicide use and agronomic practices to effectively re-duce inoculum production. To minimize the future impact of bunt, we must continue to gainan understanding of the molecular aspects of the host-parasite interaction that will permit thedesign of strategies for durable plant resistance in wheat.

Kendrick, E. L. and Holton, C. S. (1961) Racial population dynamics in Tilletia caries and T. foetida as influenced by wheat

varietal populations in the Pacific Northwest. Plant Disease Reporter 45, 5-9.


International Winter Wheat Improvement Program: BreedingModern Germplasm, Landraces and Synthetics for Disease

Resistance including Common Bunt

AlexeyMorgounov1,*, Aysel Yorgancilar2, Beyhan Akin1, Mesut Keser3, SergeyMartynov4, and Fatih Ozdemir5

1International Maize and Wheat Improvement Center (CIMMYT), Ankara, Turkey2Transitional Zone Agricultural Research Institute, Eskisehir, Turkey

3International Center for Agricultural Research in Dry Areas (ICARDA), Ankara, Turkey4Vavilov Institute, St. Petersburg, Russia

5Bahri Dagdas International Agricultural Research Institute, Konya, Turkey*[email protected]

International winter wheat improvement program (www.iwwip.org) is a partnership betweenMinistry of Food, Agriculture and Livestock of Turkey, CIMMYT and ICARDA to developnew winter wheat varieties for the region of Western and Central Asia. IWWIP also serves asa vehicle of global winter wheat exchange for the breeding program. The main breeding prio-rities are broad adaptation, yellow, leaf and stem rust resistance, grain quality. There are threemajor germplasm breeding directions: full irrigation (>5 t/ha); supplementary irrigation (2.5 –5 t/ha) and semi-arid (< 2.5 t/ha). Annually 800-1000 crosses are made which are subjectedto conventional multi-locational breeding framework in Turkey. Around 500 new varieties andbreeding lines are submitted to IWWIP by its collaborators for evaluation in Turkey and distribu-tion through the international nurseries. The best advanced lines as well as the best introducedlines are annually distributed through FAWWON (Facultative and Winter Wheat ObservationNursery) to more than 120 cooperators in around 50 countries. The program started in the 1970sand till now around 60 varieties were released in the region occupying more than 2 mln ha.

Common bunt represents high priority for IWWIP breeding across environments due to thefact that many famers in the region do not utilize certified treated seed and the disease is spreadacross the region. The crosses are made utilizing IWWIP or introduced common bunt resistancesources. Simple crosses winter x spring or winter x winter are normally top-crossed by welladapted winter germplasm preferably resistant to common bunt. The main site for selection forcommon bunt resistance is Transitional Zone Agricultural Research Institute in Eskisehir withrelatively high natural infection of common bunt. Common bunt population in the region isvirulent to Bt2, Bt3, Bt4, Bt6, Bt7, Bt12 genes based on reaction of differential set. Genotypeswith Bt1, Bt5, Bt8, Bt9, Bt10, Bt11 and Bt13 genes were resistant to pathogen. Modifiedpedigree selection scheme is applied and the susceptible populations are discarded in F2 – F3. F4

head rows are also discarded based on natural infection. Starting from F6 yield trial level (700 –900 lines) advanced lines are artificially inoculated by population of common bunt. Wheat seedsare inoculated by bunt spores using amount of 0.5 – 1% of wheat seeds tested. Material is plantedas 1 row x 1 m. For each row infected spikes and healthy spikes counted and disease incidencepercentage estimated. Genotypes with common bunt infected spikes below 5% are consideredresistant; genotypes with 5 – 10% of infected spikes are classified as moderate resistant andabove 10% - susceptible. The resistant and moderately resistant entries are again inoculated

and re-checked one more season. This systematic screening procedure resulted in identificationof 70 genotypes from IWWIP, Iran, Romania and Russia with proven resistance to commonbunt. The pedigree based diversity analysis separated them into two clusters. Contributionof several original landraces in two clusters was substantially different. This common buntresistance set has been planted and inoculated for final confirmation of resistance in Eskisehir,for seed multiplication in Edirne and at several other sites in Turkey for characterization forimportant diseases and agronomic traits. In 2014 this germplasm will be made available toglobal cooperators upon request.

In 2009 IWWIP initiated Turkey landrace inventory and within five years landraces werelocated and collected in more than 50 provinces of the country. In total, around 200 landraces(by name) were collected, evaluated and characterized following pedigree approach. Socio-economic data on these collections was also gathered from 1700 farmers. There are two mainobjectives of the landrace work: utilizing them for modern germplasm improvement and im-provement of the landraces themselves to be transferred back to farmers. In the second breedingapproach common bunt represents an important trait. Individual selection from the landracesand evaluation of the progenies for a number of traits has been conducted on a wide scale(30,000 headrows). Simple selection from diverse landraces has potential for their improve-ment. Crosses between the landraces and modern varieties targeting common bunt resistancehave been conducted as well and the F1s were either backcrossed or top-crossed by anotherlandrace. Selection for common bunt resistant progenies will start in F5 – F6.

IWWIP undertakes systematic efforts to breed winter wheat synthetics for utilization as par-ents. The potential of synthetics has been proven for a number of traits. IWWIP work startedin 2009 with the F3 populations originating from crosses between winter Durum varieties fromOdessa (Ukraine) and Ae. squarossa which were received from CIMMYT-Mexico. The pop-ulations demonstrated wide segregation and within four years pedigree method were appliedto select disease resistant agronomically more acceptable progenies. In 2013 more than 120hard-threshing synthetic lines were selected and entered multilocational testing for a number oftraits. They have been also inoculated by common bunt and there is a possibility of selection ofresistant genotypes which would diversify the genetic basis of resistance to this pathogen.

2


Mapping of Common Bunt Resistance Gene Bt9 in HexaploidWheat

Philipp Steffan1, Sven Bode Andersen1, Anna Maria Torp1, Anders Borgen2, Søren K.Rasmussen1, Erik Tybirk3, and Gunter Backes1,4,*

1University of Copenhagen, Department of Plant and Environmental Sciences, Thorvaldsensvej 40, 1871 Frederiksberg,Denmark

2Agrologica, Houvej 55, 9550 Mariager, Denmark3Nordic Seed A/S, 8464 Galten, Denmark

4University of Kassel, Department of Organic Agricultural Sciences, Nordbahnhofstrasse 1a, 37213 Witzenhausen, Germany*[email protected]

Knowledge of chromosomal locations of common bunt resistance genes in wheat is stillincomplete. Amongst those unmapped genes is also the highly effective gene designated Bt9.A double haploid population from a cross between wheat accession PI-554099 (ARS-GRINgene bank), carrying resistance gene Bt9, and the common bunt susceptible wheat cv. Cortez(Wiersum plant breeding, Netherlands) was utilized to map the common bunt resistance geneBt9 in wheat.

A number of 94 double haploid lines and their parents were evaluated for two years fortheir resistance reaction towards common bunt, both under field and greenhouse conditions.Resistance reactions were recorded as the percentage of bunt infected heads from all heads ofeach line. Results were log-transformed to fit a 1– 9 scale.

Genotyping was carried out with DArTSeq markers (Diversity Arrays Technology, Aus-tralia), resulting in 3521 polymorphic single nucleotide polymorphism markers. A consensusmap, allocating markers to linkage groups, was created based on the double haploid populationunder study in this project and two other populations from other studies, resulting in 24 linkagegroups.

Simple interval mapping was carried out using the SuperQTL program and the rqtl package ofthe R programming language, indicating a major QTL explaining 40% and 47% of the observedphenotypic variance in 2012 and 2013, respectively. No evidence for further QTL was found. Inorder to locate the linkage group containing the QTL the marker sequences were blasted againstpublic databases of wheat sequences. The linkage group was found to be located on the longarm of wheat chromosome 6D. We thus concluded that common bunt resistance gene Bt9 islocated on wheat chromosome 6DL.


Outcome of the European Tilletia Ringtest (ETR) between 2007and 2010

FabioMascher1, EmadM. Al-Maroof2, Olga Babayants3, Hermann Buerstmayr4, Philippe DuCheyron5, Veronika Dumalasova6, Blair Goates7, Marja Jalli8, Stefan Klause9, Peter

Kunz10, Zilvinas Liatukas11, Karl-JosefMuller12, Hartmut Spiess9, Susanne Vogelgsang13,andMariana Ittu14

1Agroscope IPV, 1260 Nyon, Switzerland2Sulaimani University, Bakrajo Iraq

3PBGI, 65036 Odessa, Ukraine4IFA Tulln, 3430 Tulln, Austria

5Arvalis Institut du vegetal, 78280 Guyancourt, France6CRI, 16106 Praha - Ruzyne, Czech Republik

7USDA-ARS, ID 83210 Aberdeen, USA8MTT, 31600 Jokioinen, Finland

9F & Z Dottenfelderhof, 61118 Bad Vilbel, Germany10GPK, 8634 Hombrechtikon, Switzerland

11Lithuanian Institute of Agriculture, 58344 Akademija, Lithuania12CBR Darzau, 29490 Neu Darchau, Germany

13Agroscope INH, 8056 Zürich, Switzerland14NARDI Fundulea, 915200, Calarasi, Romania

*[email protected]

Common bunt of wheat is a frequent disease in Europe and in the Middle East caused byseedborne and soilborne pathogens belonging to the genus Tilletia. The deployment of gene-tic resistances is one promising option to efficiently control the disease in an environmentallyfriendly manner. In organic breeding and in different breeding programmes in Eastern Europe,selection of resistant genotypes has been implemented for many years. Phenotyping of buntresistance is arduous as severity of the infections depends on climatic conditions, and pathogenspecies, virulence and vitality. Infection is not always successful and can vary between yearsand sites. In order to coordinate breeding efforts and to improve accuracy of resistance appraisal,the European Tilletia Ringtest (ETR) has been initiated involving several breeding and researchinstitutes. The present work provides an overview of the content and the size of the ringtest.The ringtest bases on the exchange of genetic material (wheat varieties, landraces and breed-ing lines) and a multisite testing scheme where each participants uses its confirmed infectionmethod. In parallel, in certain number of isolates the virulence spectrum has been determined.Overall 1222 reactions of individual wheat lines were determined between 2007 and 2010. Thetested wheat varieties show significant higher disease severity than the wheat breeding lines andthe wheat landraces. These latter genotypes have therefore a great potential for further breedingactivities such as gene postulations and gene cloning, determination of markers, but also for usein resistance breeding. The large virulence present in the inocula allows to conclude that themost relevant virulences were present in the ringtest.


Fifty Years of Spores: Breeding for Resistance to Dwarf Bunt ofWheat (Tilletia contraversa Kühn) in the Western US

D. J. Hole1,*, P. Tongnual2, and B. J. Goates3

1Utah State University, Department of Plants, Soils, and Climate, Logan, UT 84322 USA2Kasetsart University, Faculty of Agriculture at Kamphaeng Saen, Nakhon Pathom, 73140 Thailand (graduated)

3USDA/ARS, National Small Grains Research Facility, Aberdeen, Idaho, USA (retired)*[email protected]

Tilletia contraversa Kühn (TCK) is the causative organism for dwarf bunt, and has beenstudied due to the potential for yield losses and quality degradation from contaminated grain.

The Utah Agricultural Experiment Station in Logan, UT USA (41°45’52.66"N, 111°49’1.67"W)has selected for resistance to TCK continuously from 1923. The first cultivar released fromUAES resistant to some bunt races was “Relief”, a selection from the cross Hussar/Turkey 26.Relief was released in 1931, and cultivars resistant to TCK have been made available to growersin the Intermountain West of the USA since then. Despite a long history of virtually no commonor dwarf bunt infection in commercial production fields, susceptible cultivars trialed in thesesame farmers’ fields invariably result in a natural infection in most years.

TCK teliospores are slow to germinate and have been characterized as having a long viability[1]. Resistance to dwarf bunt in the US has largely been from germplasm collected from Turkey,and a limited number of resistance genes are known. Since new races occur infrequently, it isimportant to know viability for teliospores in the soil.

The Utah State Agricultural Experiment Station has collected spores from field infectionnurseries over sixty years. Spores had been stored as sori in intact wheat heads in ambientconditions at Logan, UT, USA, and this collection of spores was utilized to evaluate germinationin soil extract agar at 4 C under low lights [2] over an eight week period.

During the germination period, spore germination from younger collections was more rapidwith initial germination at two weeks. Some germination from spores older than twenty fiveyears was observed. The pattern of germination and differences in conidial morphology of dif-ferent collections are discussed.

1. E.J. Trione. (1982) Dwarf bunt of wheat and its importance in international wheat trade., Plant Disease, 66: 1083-1088.

2. B.J..Goates. (1996) Common and Dwarf Bunt, In: Bunt and Smut Diseases of Wheat: Concepts and Methods of Disease

Management., Wilcoxson, R.D., and E.E. Saari,D.F. (Eds.), 12-25 (CIMMYT Mexico).


Screening for Resistance to Common Bunt (Tilletia tritici) inDanish Spring and Winter Varieties of Wheat and Triticale

Bent J. Nielsen

Department of Agroecology, Flakkebjerg, Aarhus [email protected]

Susceptible varieties can suffer very serious attacks of common bunt. Fortunately, less sus-ceptible varieties exist, which only are affected to a minor degree. In order to throw light uponthe resistance situation in wheat and triticale studies testing Danish varieties were carried outin different organic projects (ORGSEED and SEED). The tests were carried out with artificialinoculation (5 g bunt spores/kg seed) in field experiments with different isolates (populations)of T. tritici (selected from 50 – 60 populations collected from different farms and productionplants in Denmark). Varieties which in previous trials had shown some resistance where testedagain on new test isolates. Most of the Danish varieties of wheat and triticale have been testeduntil 2009.

Winter and spring wheatThe studies were carried out at Aarhus University, Department of Agroecology with new

spring and winter wheat varieties. In winter wheat most varieties were – as expected – verysusceptible to attack. But some varieties suffered only minor attacks (e.g. the variety Penta)despite very severe infections in the reference varieties. The Swedish variety Stava was highlyresistant and was not attacked at all. The spring wheat varieties studied were generally alsosusceptible and only varieties such as Dragon and Leguan suffered relatively minor attacks.

In the SEED project Breeders at Nordic Seed have carried out back crossings of effectivegenes for bunt resistance into spring wheat varieties with a good general resistance to othermajor diseases. Sublines from two lines have been tested using artificial inoculation in fieldexperiments with two different isolates of T. tritici . The results clearly show a distributionbetween resistant crossing material and susceptible crossing materials.

Winter and spring TriticaleUnlike the wheat varieties most triticale varieties showed a certain degree of resistance to

bunt. Of 31 varieties studied 15 were highly resistant and suffered no attacks and 13 varietiessuffered only slight attacks (< 2% infected ears). Three of the varieties studied were susceptible(Trigantus, Triamant and He Ti 301), but even these susceptible varieties suffered less attack (8 –14% infection) than the susceptible wheat varieties. The spring triticale varieties were generallyresistant to bunt, and in the test only the variety Dublet was found to be susceptible. Some of thewinter triticale varieties were also tested for resistance to flag smut of rye (Urocystis occulta).All tested varieties were complete resistant to attack.

Potential for triticaleStudies from the organic projects shows that the majority of the winter triticale varieties

studied were resistant and that the spring triticale varieties were highly resistant. The achievedresults show an exciting potential for triticale in organic farming.


The Effect of Spore Origin when Screening for Resistance againstCommon Bunt

Anders Borgen

Agrologica, Houvej 55, 9550 Mariager, [email protected]

Screening wheat varieties for resistance against common bunt is often done by adding auniform sample of spores to a number of wheat varieties. The spore sample is often maintainedon a susceptible variety from year to year. The problem with this design is that there are noreplicates of the spore sample. It is therefore not possible to conclude what would be the resultif a different spore sample was used. A simple way to improve this system is in the second yearof experiments to test the varieties with low infection levels with both the spore sample, andin addition to that to use spores of the same variety from previous years trial. Often, the useof spores from the same variety will result in a higher infection level than using spores fromother varieties, and this indicates the build up of virulence against one or a limited number ofresistance genes. In the third year of experiments, cross inoculation of varieties with sporesfrom different semi resistant varieties can give valuable information about which varieties havethe same or different specific resistance genes.


Common Bunt and Dwarf Bunt Resistance in Winter WheatCultivars

Veronika Dumalasova

Crop Research Institute, Drnovská 507, CZ-16106 Praha-Ruzyne, Czech [email protected]

A wide range of winter wheat genotypes was tested from 1995 to 2013 for resistance tocommon bunt. The inoculum was a mixture of Czech isolates. Its virulence to Bt1, Bt2 and Bt7genes was determined on a set of differential varieties carrying individual Bt genes. Genotypeswith Bt9, Bt10, Bt11 and Bt12 genes were resistant to the inoculum mixture.

Different inocula were used occasionally, showing different reactions on the same varieties.Currently, some results indicate the presence of a broader virulence in the Czech common buntisolates. Only few of the lines that carry the Bt genes were completely free of diseased spikesin all of the tests.

In spite of the remarkable level of durable disease control that can be conferred by singleresistance genes, the genetic basis for bunt control is narrow. Wheat lines that carry knownand unknown resistance genes or new combinations of resistance genes that could be useful incultivar development are required. For this purpose a doubled haploid mapping population ofa cross between resistant variety Trintella and the susceptible variety Piko was investigated. Agenetic map was constructed using polymorphic simple sequence repeat markers and used forQTL analysis. Results indicated that resistance to common bunt could mostly be attributed to agene on chromosome 1B, a smaller QTL effects were ascribed to chromosomes 7A, 7B and 5B.

As majority of the wheat production in the Czech Republic is not conducted under organicconditions, particular attention is focused especially on dwarf bunt. Nevertheless, the prelim-inary screening was carried out with common bunt. In total, 46 of the foreign varieties testedsince 1995 in Praha-Ruzyne displayed high resistance to common bunt (no bunt incidence), 13were resistant (0-10% of spikes diseased) and some additional potential sources of resistancemay provide the screening of Czech registered varieties and the European Tilletia cooperativetest. Their reaction to dwarf bunt has to be carefully examined to specify their value for breed-ing.

Most of our dwarf bunt tests were carried out in the seedbeds with artificial inoculation.The highest infection level achieved in 2013 was 17.7%. Genotypes carrying Bt11 and Bt13genes were disease free. Up to 57.7% dwarf bunt infection was obtained in the pots with seedinoculated in the laboratory.

As there is a lack of effective genes for bunt resistance in Czech varieties with good agro-nomic adaptation, solid background for future simultaneous selection for resistance and positiveagronomic characteristics is necessary.


Mapping Dwarf Bunt Resistance in Winter Wheat – ProjectOutline and Preliminary Results

Almuth EliseMuellner and Hermann Buerstmayer*

BOKU / IFA Tulln, Institute for Biotechnology in Plant Production, Konrad-Lorenz-Str. 20, 3430 Tulln, Austria*[email protected]

Plant resistance against seed– and soil-borne diseases, such as common bunt (caused byTilletia laevis and Tilletia tritici) and dwarf bunt (caused by Tilletia controversa) is an impor-tant characteristic of varieties adopted by organic farmers. However, breeding efforts for dwarfbunt resistance in winter wheat were stopped with the advent of effective seed treatment meth-ods in the 1950s, and have not been resumed since then: Classical resistance breeding basedon phenotypic field selection is extremely cost and time consuming. In contrast, identificationof resistant lines with molecular markers could be easily integrated into existing breeding pro-grams and would be an efficient way to develop locally adapted, dwarf bunt resistant varieties –varieties which organic farmers urgently need in certain parts of Austria.

To date, the genetics of dwarf bunt resistance has been explored only marginally. Our projectaims to map dwarf bunt resistance QTL in highly resistant winter wheat genotypes and developuseful molecular markers for application in organic wheat breeding. To that end, several map-ping populations, based on crossings between dwarf bunt resistant lines (mainly old landraces)and susceptible but modern cultivars were set up. These mapping populations will be evaluatedin multi-location field trials over 3 years from 2014 to 2016 for dwarf bunt resistance. 2013served as “pilot test year” since it is quite demanding to artificially provoke severe dwarf buntinfection levels: A subset of available mapping populations was tested at 2 locations in Austria,Tulln (Lower Austria) and Lambach (Upper Austria), for dwarf bunt resistance. Artificial ino-culation was not successful in Tulln; the trial in Lambach – although dwarf bunt infection levelswere low – could be evaluated. 2014, the majority of our mapping populations will be tested infield trials at 2 locations in Austria, Schönfeld (Lower Austria) and Lambach (Upper Austria),and 1 location in Germany. Phenotypic evaluation in 2014 will lead to the identification of 2–3 promising populations which will subsequently be used for genotypic analysis, planned for2015. The combined analysis of phenotypic and genotypic data will hopefully allow us to mapdwarf bunt resistance QTL in winter wheat.

This project is funded by the Austrian Federal Ministry of Agriculture, Forestry, Environment and Water Management,

Vienna, and part of the ERA-NET project COBRA (Coordinating organic plant breeding activities for diversity): http://www.

organicresearchcentre.com/?go=Research%20and%20development&page=Plant%20breeding&i=projects.php&p_id=42

http://www.organicresearchcentre.com/?go=Research%20and%20development&page=Plant%20breeding&i=projects.php&p_id=42

http://www.organicresearchcentre.com/?go=Research%20and%20development&page=Plant%20breeding&i=projects.php&p_id=42


Study on Time Duration of the Infestation Potential of CommonBunt (Tilletia caries) and Dwarf Bunt (T. controversa) Spores of

Wheat in Soil and Farmyard Manure taking into AccountDifferent Crop Rotation Systems in Ecological Farming

R. Bauer1,*, B. Voit1, B. Killermann1, and K.-J. Hulsbergen2

1Bayerische Landesanstalt für Landwirtschaft, Vöttingerstrasse 38, 85354 Freising, Germany2TU-München, Lehrstuhl für Ökologischen Landbau und Pflanzenbausysteme, Alte Akademie 12, 85354 Freising, Germany

*[email protected]

This research work is scoping on whether in the case of huge infestation with common and/ordwarf bunt spores in soil farmers have to stop temporarily wheat cultivation and furthermorehow many years wheat should not be grown on these fields. For trying to answer these ques-tions 3-years randomized field trials are performed at 3 sites with 4 replications (10 m² per plot)on infested fields with crop rotation links commonly used in ecological farming to determinewhether it is possible to decrease the spore potential in soil. Brassica species setting free isoth-iocyanate after mulching are cultivated to examine a possible reduction of the spore viability.Additionally, the influence of stable manure on the number of bunt spores is tested.

Soil samples are taken half-yearly from each plot and common and dwarf bunt spores areisolated by wet-sieving and sedimentation steps. Spore potential is determined under the micro-scope as well as the germination ability of the spores on agar plates. Variation of the number ofspores in stable manure is determined half-yearly during storage.

Retrieval rates of about 40 % for T. controversa spores and about 25 % for T. caries sporesfrom soil could be achieved under experimental conditions. After one year storage spore poten-tial in the stable manure has decreased more than 90 %. Optimal germination conditions on agarplates for common and dwarf bunt spores from bunt balls have established. These conditionsare tested at present with spores from soil.

In addition to the crop rotation field experiment, 15 wheat varieties were tested for three yearsat two different sites in 4 replications (10 m row length) for their susceptibility to dwarf buntinfection. Bunt spore potential in soil at the test sites was of natural origin. Susceptibility of thewheat varieties was evaluated by counting the number of infected ears per row and number ofspores per kernel.

Three groups could be distinguished: a highly susceptible, an intermediate, and a rathertolerant group, whereas the first and the last one could be statistically distinguished.


Phosphatases from Ustilago maydis and Possible Role duringInfection

Trine H. Sørensen1, Lars K. Skov1, and Søren K. Rasmussen2,*

1Novozymes A/S, Copenhagen, Denmark2University of Copenhagen, Department of Plant and Environmental Sciences, Thorvaldsensvej 40, 1871 Frederiksberg,

Denmark*[email protected]

Three genes encoding putative phytases (um03382, um04282 and um06428) have been iden-tified in the biotrophic pathogen Ustilago maydis. These three phytases contain conservedamino acid residues typically class of histidine acid phosphatases. Two of the putative phy-tase encoding genes (um03382 and um04282) isolated from U. maydis were heterologous ex-pressed in Aspergillus oryzae. It has been established that the um04282 encoded protein isa general phosphatase and the um03382 encoded protein has phytase activity. Hence theyare denominated Umphos and Umphy. The pH and temperature profile of Umphy resemblesthat of the other characterized phytases of the histidine acid phosphatases family. Isomer-specific high-pressure liquid chromatography analysis, 1H and 31P NMR established myo-inositol(1,2,4,5,6)pentapkishosphate as the major product formed when InsP6 is dephospho-rylated by Umphy.

In many aspects U. maydis is regarded as a model organism for elucidating basic cell biologicprocesses and biotrophic interaction (Brefort et al. 2009). The intuitive biological relevance ofUmphy and Umphos in Ustilago maydis is the role of these enzymes in phosphate acquisition.Potentially the Umphos and Umphy could be regulated by changes in phosphate level, which isseen for other genes in U. maydis involved in phosphate metabolism (Larraya et al. 2005).

Potentially, the biological relevance of Umphy in U. maydis could be to acquire phosphatein a controlled manner where only one phosphate group is gained from the myo-inositol ring.This strategy of only “stealing” one phosphate from the plant could be in accordance with thebiotrophic lifestyle.

As phosphorylated myo-inositols are ubiquitously found in all cells and have been associatedwith numerous signaling pathways, it is reasonable to consider the possibility of Umphy beinginvolved in formation of secondary messengers. As Umphy are extracellular expressed andmyo-inositol and its phosphorylated derivatives have been associated with various aspects ofplant defense, the specific hydrolysis products of InsP6 by Umphy could contribute to inhibitionof plant defense, which would be in accordance with the biotrophic lifestyle of U. maydis.


Common Bunt Caused by Tilletia caries: Evaluation of ViableSpores, Set Up of a Protocol to Access Transmission to Plantlets

and Damage Threshold

G. Orgeur1,*, A. Delaunay1, I. Serandat1, F. Decugis1, M. Rolland1, J. Gombert2, R. Valade3,and V. Grimault1

1GEVES, 25 rue Georges Morel, CS 90024, 49071 Beaucouzé, France2FNAMS, Impasse du Verger, 49800 Brain sur l’Authion, France

3ARVALIS, Domaine AgroParisTech, Bât. INRA Bioger-cpp, 78850 THIVERVAL-GRIGNON, France*[email protected]

Common bunt is a re-emerging disease due to the development of the organic farming whichuses seeds without chemical treatment (Vanova et al., 2006). Development of new alternativemethods of treatment allow to kill but do not supress the spores.

A first study was aimed to evaluate the viability of spores by diffrent ways (germination andcoloration) and to confirm interest to detect viable spores to dead spores. Seeds naturally andartificially infected (adapted to CEB method N°42) with different contamination rates of viablespores (10 000 and 1 000 spores per seed) and no viable spores (10 000 spores per seed) havebeen sown in order to evaluate the impact of viable spores of T. caries on symptoms expressionon ears the next years. The results showed that the only seeds able to induce symptoms in earsthe next year were seeds naturrally and artificially contaminated by viables spores (10 000 and1 000 spores per seed).

A second study funded by the TESTA European project aimed at defining a protocol to studydamage threshold and transmission of T. caries from seed to plantlet. Current protocols to studytransmission of T. caries use naturally or artificially contaminated seeds sown in the field andnotation of bunted ears, which are long and time consuming. The aim of our study was todevelop a protocol which was not dependant of environmental condition, not time consumingand could give an early result. Artificial seed contamination (adapted to CEB method N°42)with different contamination rates of viable spores (10 000, 1 000, 100, 1 spores per seed) andno viable spores (10 000 spores per seed) were done to evaluate the threshold damage of T.caries. Infected seeds have been sown in climatic chamber according to Eibel et al. in 2005and plants have been transferred in field and greenhouse until symptoms expression. Artificialcontamination transposed in field and greenhouse has shown that a low contamination rate of T.caries, was able to induce symptoms in ears the next year. In greenhouse, first bunt ears wereoberved at 100 spores per seeds. In field, bunt ears were observed in plants infected by 1 sporeper seeds. But symptoms were also observed in healthy seeds due to cross contamination. Thisstudy allowed to obtain a protocol of transmission from seed to planted in optimal conditions. APCR protocol was developped in order to allow early detection of Tilletia spp on plantlets. PCRdetection at cotyledon and 2 leaves stage was compared to symptom expression and the resultshave shown that detection was better in stem six weeks after sowing. Correlation between PCRdetection and symptom expression will be presented to show the efficiency of the protocol tostudy transmission of T. caries from seed to plantlets and its usefulness to access efficiency ofseed treatments, in a faster way than the actual protocol in field.


Identification of New Sources of High Levels of Resistance toDwarf Bunt and Common Bunt among Winter Wheat Landraces

in the USDA-ARS National Small Grains Collection

Blair J. Goates1,2

1United States Department of Agriculture, Agricultural Research Service, Aberdeen, Idaho, USA2Current address: 8388 N Parks RD, Pocatello, Idaho 83201, USA

[email protected]

Even though chemical seed treatments are available for control of common bunt of wheat(caused by Tilletia caries and Tilletia foetida) and dwarf bunt of wheat (caused by Tilletia con-traversa), host resistance is still an important part of cultivar development in many areas of theworld. In addition, host resistance has gained interest due to the increase in organic produc-tion. The pathogens causing these two diseases are very closely related and are controlled bythe same set of resistance genes. Few resistance genes are utilized in cultivars creating vul-nerability to previously unidentified pathogenic races that can emerge. In an attempt to findnew sources of resistance, thousands of hexaploid landrace wheats from the USDA-ARS Na-tional Small Grains Collection were screened for resistance to these diseases. Fifty-five wheataccessions that showed high levels of resistance to a composite of pathogenic races of dwarfbunt or common bunt fungi in repeated field tests under high disease pressure were tested to 6pathogenic races of common bunt fungi and 3 races of dwarf bunt fungi. These pathogenic raceshave specific virulence characteristics which are particularly useful for elucidating resistancefactors in wheat. All of the previously known sources of useful bunt resistance and most of theother lines tested were susceptible to one or more of the individual bunt races. Wheats that hadthe same or similar reactions to the races were separated into groups where common resistancegenes or gene combinations were indicated. Seven new lines were identified that were highlyresistant or immune to all of the races indicating the presence of unknown resistance genes orgene combinations. Unexpected resistant and/or susceptible reactions in particular wheat geno-type/bunt race interactions indicated the presence of unknown resistance genes in some of thewheat lines, and also indicated the presence of unknown virulence factors in the bunt races used.The identification of various common resistance factors among these resistant lines enables theselection of diverse sources of resistance to be used as parents in breeding programs. The sevennew lines that had a high level of resistance to all the bunt races should be valuable for theintrogression of new sources of resistance, broadening the genetic diversity of bunt resistancein cultivars. Whether the seven wheats have the same or different resistant factors needs to bedetermined.


Why Organic Agriculture Doesn’t Want to Use Fungicides

Gunter Backes

University of Kassel, Faculty of Organic Agricultural Sciences, Section of Organic Plant Breeding and Agrobiodiversity,Nordbahnhofstr. 1a, 37213 Witzenhausen, Germany

[email protected]

The International Federation of Organic Agriculture (IFOAM) defines Organic Agriculture(OA) as “(...) a production system that sustains the health of soils, ecosystems and people.(...)”(IFOAM 2008a), implementing that OA is defined through its process, not its result. There-fore, it is important that every part of the process follows the principles of OA, which are theprinciples of “Health”, “Ecology”, “Fairness” and “Care” as defined by IFOAM (2008b). Theapplication of (synthetic) fungicides practically violates each single of these principals, but es-pecially “Health” that comprises the wholeness and integrity of living systems and the principleof “Ecology”. Ideally in OA, the aim is to use different approaches in order to strengthen thesystem do deal with threats such as fungal epidemics, that is to work with the system (whichalso includes pathogens). Fungicides work wittingly against a part of the system and mostlyunintentionally against further parts.

Alternative approaches in OA to control fungal pathogens are (a) a wide and varied rotationthat not only reduces accumulation of pathogens but also can actively reduce their occurrence(Mazzola and Gu 2002); (b) the stimulation of high biological activity especially in the soil(Sharma et al. 2012); (c) a higher diversity in the plant population which counteracts diseaseepidemics both by spatial effects and by induced resistance through avirulent pathotypes of thepathogen mixture (Finckh et al. 2000); (d) a genetic resistance level in the cultivated (mixtureof) genotypes that effectively suppresses disease epidemics (Wächter et al. 2007); (e) as ultimaratio and especially in seed production the application of natural substances, e.g. glucosinolatesfrom mustard or milk powder against bunt (Borgen 2003) or physical treatments of the seedse.g. hot water treatments or brushing against bunts (Borgen 2005).

Borgen A (2003) Effect of seed treatment with milk powder and mustard flour in control of common bunt (Tilletia tritici) inwheat and stem smut (Urocystis occulta) in rye. Proc. from BCPC Symp. No. 76 “Seed Treat. Challenges Oppor.Borgen A (2005) Removal of bunt spores from wheat seed lots by brush cleaning. 13–15.Finckh MR, Gacek ES, Goyeau H, et al. (2000) Cereal variety and species mixtures in practice, with emphasis on diseaseresistance. Agronomie 20:813–837IFOAM (2008a) Definition of Organic Agriculture. 1IFOAM (2008b) Principles of Organic Agriculture. 4Mazzola M, Gu Y-H (2002) Wheat Genotype-Specific Induction of Soil Microbial Communities Suppressive to Disease Incitedby Rhizoctonia solani Anastomosis Group (AG)-5 and AG-8. Phytopathology 92:1300–7. doi: 10.1094/PHYTO.2002.92.12.1300Sharma K, Bruns C, Butz AF, Finckh MR (2012) Effects of fertilizers and plant strengtheners on the susceptibility of tomatoesto single and mixed isolates of Phytophthora infestans. Eur J Plant Pathol 133:739–751

Wächter R, Waldow F, Müller K-J, et al. (2007) Resistance of winter wheat varieties and breeding lines against common bunt

(Tilletia tritici) and dwarf bunt (T. controversa). Nachrichtenblatt Dtsch Planzenschutzd 59:30–39.

The XVIII Biennial International Workshop on the Smuts and ...

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